Method of treating bacterial infections using gemifloxacin or a salt thereof and a carbapenem antibacterial agent

ABSTRACT

The present invention relates to a method of treating anti-bacterial infections which method comprises the separate, simultaneous or sequential administration to a patient in need thereof, of an effective amount of gemifloxacin or a salt thereof and a carbapenem antibacterial.

TECHNICAL FIELD

This invention relates to a novel method of treating bacterialinfections using a combination of antibacterial agents, moreparticularly gemifloxacin or a salt thereof and a carbapenemantibacterial agent.

BACKGROUND ART

Gemifloxacin[(R,S)-7-(3-aminomethyl-4-methoxyiminopyrrolidin-1-yl)-1-cyclopropyl-6-fluoro-4-oxo-1,4-dihydro-1,8-naphthyridine-3-carboxylicacid] is a fluoroquinolone antibacterial agent which has enhanced invitro antibacterial activity against Gram-positive bacteria, whilstretaining excellent activity against Gram negative bacteria. Withoutintending to be bound or limited by theory, gemifloxacin is believed toact via inhibition of bacterial topoisomerase II and IV. Gemifloxacin ishighly selective for bacterial rather than human topoisomerase II.

EP 688772 discloses novel naphthyridine carboxylic acid derivatives,including gemifloxacin. WO 98/42705 discloses gemifloxacin mesylate andhydrates thereof including the sesquihydrate.

There still remains the need for improved methods of treating bacterialinfections. In particular, infections caused by Pseudomonas aeruginosacontinue to pose a therapeutic problem. In clinical practice, thecombination of β-lactam antibiotic and aminoglycoside antibacterialagents has been shown to have an improved efficacy for the treatment ofinfections caused by P. aeruginosa. However, increased resistance of P.aeruginosa to aminoglycosides, coupled with their potential fornephrotoxicity, means there still remains the need for alternativetreatments.

DISCLOSURE OF THE INVENTION

The present invention provides a method of treating bacterial infectionswhich method comprises the separate, simultaneous or sequentialadministration to a patient in need thereof, of an effective amount ofgemifloxacin or a salt thereof and a carbapenem antibacterial. Thepatient may be human or animal, and in a preferred embodiment is human.

The present inventors have found combinations of gemifloxacin or a saltthereof and a carbapenem antibacterial that provide an antibacterialregimen which has a broader spectrum of activity than either agentalone. In particular, the present invention includes such combinationshaving synergistic activity against several clinical isolates orreference strains of Pseudomonas aeruginosa, relative to either agentalone.

Suitable salts of gemifloxacin include those described in WO 98/42705,EP 688772, and U.S. Pat. No. 5,776,944. In particular embodiments, thesalt of gemifloxacin is selected from the mesylate and hydrates thereof,in particular the sesquihydrate as described in WO 98/42705.

Suitable carbapenem antibacterials for use in the method of theinvention are well known in the art and include, e.g., biapenem,imipenem, meropenem and panipenem.

In particular embodiments, the carbapenem antibacterial for use in themethod of the invention is selected from meropenem and imipenem, inparticular meropenem.

The carbapenem antibacterials referred to herein may be in the form ofthe free acids or pharmaceutically acceptable salts or in-vivohydrolysable esters.

Gemifloxacin or a salt thereof and a carbapenem antibiotic, or acomposition comprising same, may be used in accordance with the presentinvention to modulate metabolism of bacteria (e.g., clinical isolates,reference bacteria, pathogenic bacteria) and/or to treat infectionscaused by such bacteria. Clinical isolates or reference bacteria includeStreptococcus pneumoniae (e.g., ATCC 49619), Haemophilus influenzae(e.g., ATCC 49247), Moraxella catarrahalis (e.g., 1502), Staphylococcusaureus (e.g., ATCC 29213), Staphylococcus saprophyticus (e.g., 662),Klebsiella pneumoniae (e.g. E70), Proteus vulgaris (e.g., ATCC 13315),Enterococcus faecalis (e.g., ATCC 29212), Escherichia coli (e.g., ATCC25922), and Pseudomonas aeruginosa (e.g., ATCC 27853, 6016, 6156, 6168,P003, 6140, and PA018R).

Particular embodiments of the invention include the following treatmentregimens:

-   a) gemifloxacin and meropenem for the treatment of infections caused    by Pseudomonas aeruginosa; and-   b) gemifloxacin and imipenem for the treatment of infections caused    by Pseudomonas aeruginosa.

Suitably, gemifloxacin or a salt thereof and the carbapenemantibacterial are administered in a ratio of from about 10:1 to about1:10, more suitably about 5:1 to 1:5, typically about 2:1 w/w, expressedas the weight of the free acid and free base respectively.

Suitably, the administration is substantially simultaneous. This mayconveniently be achieved by the co-administration of separatepharmaceutical compositions comprising gemifloxacin or a salt thereofand a carbapenem antibacterial. Such separate compositions may beusefully provided as a kit comprising a gemifloxacin, or a salt thereof,composition and a carbapenem antibacterial composition. The kitpreferably contains sufficient dosages of gemifloxacin and thecarbapenem antibacterial for a single course of therapy for theparticular infection to be treated, together with instructions foradministration.

Accordingly the present invention also provides a kit of parts for usein treating bacterial infections in mammals which comprises anantibacterially effective amount of (a) a pharmaceutical compositioncomprising gemifloxacin or a salt thereof, and a pharmaceuticallyacceptable carrier, and (b) a pharmaceutical composition comprising acarbapenem antibacterial and a pharmaceutically acceptable carrier.

Alternatively gemifloxacin, or a salt thereof, and a carbapenemantibacterial may be formulated together and administered in a singlecomposition.

Accordingly in a further aspect the present invention further provides apharmaceutical composition comprising gemifloxacin or a salt thereof, acarbapenem antibacterial, and a pharmaceutically acceptable carrier.

The present invention also includes the use of gemifloxacin or a saltthereof in the manufacture of a medicament for use in combination with acarbapenem antibacterial in the treatment of bacterial infections.

The invention further provides gemifloxacin or a salt thereof incombination with a carbapenem antibacterial for use in the treatment ofbacterial infections.

The invention also includes a method of treating bacterial infectionscomprising administering to a mammal in need of such treatment, atherapeutically effective amount of gemifloxacin or a salt thereof and acarbapenem antibacterial. In preferred embodiments the treatmentcomprises administering therapeutically effective amounts ofgemifloxacin or a salt thereof and a carbapenem antibiotic, wherein theactivity of the antibiotics againt the bacterial infection issynergistic.

The invention also includes a method of modulating the metabolism ofbacteria, comprising contacting the bacteria with an antibacteriallyeffective amount of gemifloxacin and a salt thereof and a carbapenemantibiotic (optionally in the form of an antibacterially effectivecomposition or kit of compositions, as described herein). Modulatingmetabolism suitably comprises inhibiting growth of the bacteria orkilling the bacteria. Contacting the bacteria may suitably comprise thestep of introducing the antibiotics or composition or kit ofcompositions comprising same into a mammal. In preferred embodiments,the method comprises contacting the bacteria with an antibacteriallyeffective amount of gemifloxacin or a salt thereof and a carbapenemantibiotic, or composition or kit of compostions comprising same,wherein the activity of the antibiotics against the bacteria issynergistic.

Suitable formulations comprising gemifloxacin include those described inWO 98/42705, EP 688772, U.S. Pat. No. 5,776,944.

Suitable formulations comprising a carbapenem antibacterial are wellknown in the art and are readily available commercially.

Gemifloxacin or a salt thereof may be formulated with a carbapenemantibacterial and standard pharmaceutical carriers or diluents accordingto conventional procedures well known in the art. These procedures mayinvolve mixing, granulating and compressing or dissolving theingredients as appropriate to the desired preparation.

The invention further provides a method for the preparation of apharmaceutical composition comprising gemifloxacin or a salt thereof anda carbapenem antibacterial which method comprises admixing thecombination of gemifloxacin or a salt thereof and a carbapenemantibacterial and a pharmaceutically acceptable carrier.

The invention further provides a formulation for the treatment ofantibacterial infections comprising gemifloxacin or a salt thereof and acarbapenem antibacterial. In preferred embodiments, the formulationcomprises an amount of gemifloxacin or a salt thereof and carbapenemantibiotic wherein the activity of the antibiotics in treating thebacterial infection is synergistic.

The invention further provides the use of a formulation comprisinggemifloxacin or a salt thereof and a carbapenem antibacterial in themanufacture of a medicament for the treatment of bacterial infections.

In the compositions, kits, and methods of the present invention thebacterial infection is preferably one caused by P. aeruginosa.

Infections caused by P. aeruginosa include wound infections, urinarytract infections and respiratory tract infections, together with generalinfections in an immunocompromised patient.

The contacting step and administration step in any of the methods of theinvention may be performed in many ways that will be readily apparent tothe skilled artisan. However, it is preferred that the contacting stepand administration step is a provision of a composition comprisinggemifloxacin or a salt thereof and a carbapenem antibiotic, or in thecase of a kit according to the present invention, a compositioncomprising gemifloxacin or a salt thereof and a composition comprising acarbapenem antibiotic, to a human patient in need of suchcomposition(s), or directly to bacteria in culture medium or buffer.

For example, when contacting a human patient or contacting said bacteriain a human patient or in vitro, the compositions or kit of compositionscomprising gemifloxacin or a salt thereof and a carbapenem antibiotic,preferably pharmaceutical compositions, may be administered in anyeffective, convenient manner including, for instance, administration bytopical, oral, anal, vaginal, intravenous, intraperitoneal,intramuscular, subcutaneous, intranasal or intradermal routes amongothers.

It is also preferred that these compositions be employed in combinationwith a non-sterile or sterile carrier or carriers for use with cells,tissues or organisms, such as a pharmaceutical carrier suitable foradministration to a subject. Such compositions comprise, for instance, atherapeutically effective amount of gemifloxacin or a salt thereof,and/or a carbapenem antibiotic, a pharmaceutically acceptable carrier orexcipient and optionally a media additive. Such carriers may include,but are not limited to, saline, buffered saline, dextrose, water,glycerol, ethanol and combinations thereof. The formulation should suitthe mode of administration.

In therapy or as a prophylactic, the gemifloxacin or salt thereof andcarbapenem antibiotic are preferably administered to an individual as aninjectable composition (including compositions comprising bothantibiotics, or separate injectable compositions comprising one or theother antibiotic), for example as a sterile aqueous dispersion,preferably an isotonic one.

Alternatively, the gemifloxacin or salt thereof and carbapenemantibiotic in the methods of the invention may be formulated for topicalapplication for example in the form of ointments, creams, lotions, eyeointments, eye drops, ear drops, mouthwash, impregnated dressings andsutures and aerosols, and may contain appropriate conventionaladditives, including, for example, preservatives, solvents to assistdrug penetration, and emollients in ointments and creams. Such topicalformulations may also contain compatible conventional carriers, forexample cream or ointment bases, and ethanol or oleyl alcohol forlotions. Such carriers may constitute from about 1% to about 98% byweight of the formulation; more usually they will constitute up to about80% by weight of the formulation.

For administration to mammals, and particularly humans, it is expectedthat the antibacterially effective amount is a daily dosage level of theactive agent from 0.001 mg/kg to 10 mg/kg, typically around 0.1 mg/kg to1 mg/kg, preferably about 1 mg/kg. A physician, in any event, willdetermine an actual dosage that is most suitable for an individual andwill vary with the age, weight and response of the particularindividual. The above dosages are exemplary of the average case. Therecan, of course, be individual instances where higher or lower dosageranges are merited, and such are within the scope of this invention. Itis preferred that the dosage is selected to modulate metabolism of thebacteria in such a way as to inhibit or stop growth of said bacteria orby killing said bacteria. The skilled artisan may identify this amountas provided herein as well as using other methods known in the art, e.g.by the application MIC tests.

A further embodiment of the invention provides for the contacting stepor administration step of the methods to further comprise contacting anin-dwelling device in a patient. In-dwelling devices include, but arenot limited to, surgical implants, prosthetic devices and catheters,i.e., devices that are introduced to the body of an individual andremain in position for an extended time. Such devices include, forexample, artificial joints, heart valves, pacemakers, vascular grafts,vascular catheters, cerebrospinal fluid shunts, urinary catheters, andcontinuous ambulatory peritoneal dialysis (CAPD) catheters.

The gemifloxacin or salt thereof and carbapenem antibiotic, orcomposition or kit of compositions of the invention, may be administeredby injection to achieve a systemic effect against relevant bacteria,shortly before insertion of an in-dwelling device. Treatment may becontinued after surgery during the in-body time of the device. Inaddition, the composition or kit of compositions could also be used tobroaden perioperative cover for any surgical technique to preventbacterial wound infections.

In addition to the therapy described above, gemifloxacin or a saltthereof and a carbapenem antibiotic, or composition or kit ofcompositions used in the methods of this invention may be used generallyas a wound treatment agent to prevent adhesion of bacteria to matrixproteins, exposed in wound tissue and for prophylactic use in dentaltreatment as an alternative to, or in conjunction with, antibioticprophylaxis.

Alternatively, gemifloxacin or a salt thereof and a carbapenemantibiotic, or composition or kit of compositions of the invention maybe used to bathe an indwelling device immediately before insertion. Theactive agent will preferably be present at a concentration of 1 μg/ml to10 mg/ml for bathing of wounds or indwelling devices.

All documents cited or referred to herein, including issued patents,published and unpublished patent applications, and other publicationsare hereby incorporated herein by reference as though fully set forth.

Best Mode for Carrying out the Invention

The invention will now be described by the following examples which areillustrative and not intended to limit the invention hereinbeforedescribed.

EXAMPLES

The potential for the combination therapy of gemifloxacin and acarbapenem antibacterial was investigated.

1) Gemifloxacin and Meropenem

The combined effect of gemifloxacin and meropenem was investigated in P.aeruginosa by using a checkerboard titration method. Evaluation forsynergistic activity was carried out by calculation of fractionalinhibitory concentrations (FICs) according to the method of Eliopouloset al (1996, Antimicrobial Combinations, Antibiotics in LaboratoryMedicine, Victor Lorian, 4th Edition, pp 337-338, Baltimore, Md.,Williams and Wilkins).

P. aeruginosa ATCC 27853 was obtained from the SmithKline BeechamAnti-Infectives Research Culture Collection and stored at −80° C. in 10%glycerol. Before testing, the isolate was passaged onto agar plates(trypticase soy agar containing 5% sheeps blood) from the frozen stockfor two consecutive days. Cation adjusted Mueller Hinton broth (BBL,Cockeyville, Md.) was used for the isolate.

Checkerboard microtitre broth dilution plates were prepared using theHamilton MicroLab AT Plus system (Reno, Nev.). Two-fold serial dilutions(50 μl) of meropenem were made in columns 1 through 11 of a microtitreplate. Two-fold serial dilutions of gemifloxicin were prepared manuallyand the MicroLab AT Plus was used to dispense 25 μl of gemifloxacin ateach concentration into rows A through G of the microtitre plate. Theconcentration range tested varied for each drug/organism combination toencompass the MIC endpoints of the individual components. Column 12contained two fold serial dilutions of gemifloxacin only, and row Hcontained two fold serial dilutions of meropenem only. The last well (H)in column 12 was used as a positive growth control well, containing onlymedium and test isolate.

The bacteria were diluted to a 0.5 McFarland standard and then furtherdiluted {fraction (1/50)}. Each well was inoculated with 25 μl of theisolate to give a final inoculum density of approximately 5×10⁵ cfu/ml.The MicroLab AT Plus 2 was used to add the inoculum to the microtitreplates. After inoculation, the plates were covered with a 96 wellmicrotitre plate lid and incubated at 35° C. in ambient air for 20-24hours. A 10 μl aliquot of the inoculum was plated on trypticase soy agarcontaining 5% sheep blood to determine the purity of the final testinoculum.

Following incubation, a microtitre mirror reader (Cooke InstrumentsLtd., England) was used to assist in determining the microdilution MICendpoints. The MIC was determined as the lowest concentration ofcompound that inhibited visible growth of the organism.

The Fractional Inhibitory Concentrations (FICs) were calculated usingthe following formula:(A)/(MIC A)+(B)/(MIC B)=FIC A+FIC B=FIC Indexwherein:

-   A is the MIC of drug A in the presence of drug B-   B is the MIC of drug B in the presence of drug A-   MIC A is the MIC of the organism to drug A alone-   MIC B is the MIC of the organism to drug B alone-   FIC A is the fractional inhibitory concentration of drug A-   FIC B is the fractional inhibitory concentration of drug B

The FIC indices were interpreted using the following criteria:

-   ≦0.5=Synergy-   >0.5-1=Additive-   >1-2=Indifference-   >2=Antagonism

Synergy was observed against P. aeruginosa 27853 for thegemifloxacin/meropenem combination. gemifloxacin μg/ml 0.125 — meropenemμg/ml — 0.125 gemifloxacin MIC μg/ml — 0.125 meropenem MIC μg/ml 0.125 —FIC 0.5  0.5  gemifloxacin MIC 0.5  0.5  meropenem MIC 0.5  0.5 

Other combinations of gemifloxacin/meropenem tested in a similar mannerexhibited an additive or indifferent effect against P. aeruginosa 27853.In wells where no growth was observed and the MIC of one of the agentshad been achieved or exceeded, the result was reported as inhibited.

2. Gemifloxacin and Imipenem

The combined effect of gemifloxacin and imipenem was investigated in P.aeruginosa by using a checkerboard titration method. The test organismsused in this study were all clinical isolates collected in Korea. Theisolates were maintained frozen at −70° C. prior to testing.

Gemifloxacin was tested at 12 concentrations (0.016-32 μg/ml) andimipenem was tested at 11 concentrations (0.016-16 μg/ml). Gemifloxacinwas dispensed alone in the first row and combined with imipenem in theremaining rows. Imipenem solution was also dispensed alone in the firstrow in the first column. Test strains were grown for 18 h inMueller-Hinton Broth, and then these overnight cultures were dilutedwith the same fresh medium to the density of approximately 10⁷ CFU/mL.The bacterial concentrations were determined by measuring opticaldensity or turbidity of the suspension and were verified by standardcolony counts on antibiotic-free agar plates. A diluted bacterialsolution was applied to plates containing serially diluted antimicrobialagents to yield 5×10⁵ CFU/mL. Plates were incubated at 35° C. for 18hours.

Minimal inhibitory concentrations (MICs) for each separate drug weredetermined. For wells along the growth-no growth interface, synergy wasdetermined by calculating the Fractional Inhibitory Concentration (FIC)index. The Fractional Inhibitory Concentration (FIC) indices werecalculated using the following formula:FIC index=FIC _(A) +FIC _(B) =[A]/MIC _(A) +[B]/MIC _(B)Wherein:

-   FIC_(A(or B)), is the FIC of drug A (or B)-   MIC_(A (or B)) is the MIC of the organism to drug A (or B)-   A (or B) is concentration of drug A (or B) that is the lowest    inhibitory concentration

The FIC indices were interpreted using the following criteria:

-   ≦0.5=Synergy;-   >0.5-1.0=Additive;-   >1.0-4.0=Indifference;-   >4.0=Antagonism

Synergy against P. aeruginosa 6168, 6140 and PA018R was observed forcombinations of gemifloxacin and imipenem. P. gemifloxacin imipenemC_(gemifloxacin) C_(imipenem) aeruginosa MIC μg/ml MIC μg/ml MIC μg/mlMIC μg/ml FIC_(A) FIC_(B) 6168 0.5 1 0.0625 0.25 0.13 0.25 6140 8 4 2 10.25 0.25 PA018R 4 16 1 1 0.25 0.06

Combinations of gemifloxacin and imipenem tested in a similar mannerexhibited an additive effect against P. aeruginosa 6016, 6156, and P003.

INDUSTRIAL APPLICABILITY

According to the present invention, combinations of gemifloxacin or asalt thereof and a carbapenem antibacterial provide an antibacterialregimen which has a broader spectrum of activity than either agentalone. In particular, such combinations has synergistic activity againstseveral clinical isolates or reference strains of Pseudomonasaeruginosa, relative to either agent alone.

1. A method of treating anti-bacterial infections which method comprises the separate, simultaneous or sequential administration to a patient in need thereof, of an effective amount of gemifloxacin or a salt thereof and a carbapenem antibacterial.
 2. A method according to claim 1 comprising administration of gemifloxacin mesylate.
 3. A method according to claim 1 wherein the carbapenem antibacterial is meropenem or imipenem.
 4. A method according to claim 3 wherein the carbapenem antibacterial is meropenem.
 5. A method according to claim 1 wherein gemifloxacin or a salt thereof and a carbapenem antibacterial are administered in a ratio of from about 10:1 to about 1:10 (w/w).
 6. A method according to claim 1 wherein the bacterial infection is caused by Pseudomonas aeruginosa.
 7. A method according to claim 1 in which an effective amount of gemifloxacin or a salt thereof and a carbapenem antibacterial are administered substantially simultaneously.
 8. A method according to claim 6 achieved by the co-administration of separate compositions comprising gemifloxacin or a salt thereof and a carbapenem antibacterial.
 9. A kit comprising a gemifloxacin, or a salt thereof, formulation and a carbapenem antibacterial formulation.
 10. A method according to claim 2 wherein the carbapenem antibacterial is meropenem or imipenem.
 11. A method according to claim 10 wherein the carbapenem antibacterial is meropenem. 